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Abstract DSN (duplex-specific nuclease ) displays a strong preference for cleaving ds DNA and DNA in DNA–RNA hybrid duplexes compared with ss DNA and RNA irrespective of sequence length , as described recently. The technique termed DSN is a simple and effective cDNA normalization method. In order to screem and isolate new and full-length sporulation-specific genes of entomopathogenic fungus Metarhizium in the sporulation stage with great efficiency, a combined DSN normalization method with the SMARTTM (switching mechanism at 5′end of RNA transcript) library construction method was used to construct a normalization and full-length cDNA library of entomopathogenic fungus Metarhizium in the sporulation stage. The titer of unamplified cDNA library was 2.1 ×106 cfu/mL and contained 6×106 independent clones.The average cDNA inserts size was 1500bp with a recombination rate of 95%.Sequencing results and bioinformatics analysis of random picked clones indicated that the ratio of full-length cDNA was about 60％.The presentation levels of the abundant transcripts G3PDH and β-tubulin decreased obviously compared with non-normalized samples.
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Received: 09 January 2007
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